title: Actin polymerization does not provide direct mechanical forces for vesicle fission during clathrin-mediated endocytosis.
abstract: Actin polymerization is important for vesicle fission during clathrin-mediated endocytosis (CME), and it has been proposed that actin polymerization may promote vesicle fission during CME by providing direct mechanical forces. However, there is no direct evidence in support of this hypothesis. In the present study, the role of actin polymerization in vesicle fission was tested by analyzing the kinetics of the endocytic tubular membrane neck (the fission-pore) with cell-attached capacitance measurements to detect CME of single vesicles in a millisecond time resolution in mouse chromaffin cells. Inhibition in dynamin GTPase activity increased the fission-pore conductance (Gp), supporting the mechanical role of dynamin GTPase in vesicle fission. However, disruptions in actin polymerization did not alter the fission-pore conductance Gp, thus arguing against the force-generating role of actin polymerization in vesicle fission during CME. Similar to disruptions of actin polymerization, cholesterol depletion results in an increase in the fission-pore duration, indicating a role for cholesterol-dependent membrane reorganization in vesicle fission. Further experiments suggested that actin polymerization and cholesterol might function in vesicle fission during CME in the same pathway. Our results thus support a model in which actin polymerization promotes vesicle fission during CME by inducing cholesterol-dependent membrane reorganization.
authors.affiliation: Departments of Biological Sciences, Bioengineering, and Chemistry, University of Illinois at Chicago, Chicago, Illinois 60607.
authors.forename: Li-Hua
authors.initials: LH
authors.lastname: Yao
authors.forename: Yan
authors.initials: Y
authors.lastname: Rao
authors.forename: Chi
authors.initials: C
authors.lastname: Bang
authors.forename: Svetlana
authors.initials: S
authors.lastname: Kurilova
authors.forename: Kelly
authors.initials: K
authors.lastname: Varga
authors.forename: Chun-Yang
authors.initials: CY
authors.lastname: Wang
authors.forename: Brandon D
authors.initials: BD
authors.lastname: Weller
authors.forename: Wonhwa
authors.initials: W
authors.lastname: Cho
authors.forename: Jun
authors.initials: J
authors.lastname: Cheng
authors.forename: Liang-Wei
authors.initials: LW
authors.lastname: Gong
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D019439:Chromaffin Cells / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism*; D034281:Dynamins / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism*; D051379:Mice; D058105:Polymerization
chemical_list: D000199:Actins; D002966:Clathrin; D034281:Dynamins